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Abstract:

Disclosed is a process comprising the steps of: expressing juice from
fresh tea leaves thereby to produce leaf residue and juice comprising a
mixture of tea compounds; fractionating the mixture; and recovering at
least one fraction enriched in at least one tea compound.

Claims:

1. A process comprising the steps of:a) expressing juice from fresh tea
leaves thereby to produce leaf residue and juice comprising a mixture of
tea compounds;b) fractionating the mixture; andc) recovering at least one
fraction enriched in at least one tea compound.

9. A process according to claim 8 wherein the leaf tea is black leaf tea.

10. A process according to claim 1 wherein the moisture content of the
fresh tea leaves from which juice is expressed in step (a) is from 30 to
90% by weight of the fresh tea leaves.

11. A process according to claim 1 wherein the mixture of tea compounds is
not diluted with further tea compounds prior to or during step (b).

12. A process according to claim 11 wherein the juice is not mixed with
tea extract prior to or during step (b).

Description:

TECHNICAL FIELD OF THE INVENTION

[0001]The present invention relates to a process for purifying compounds
from tea. More particularly the present invention relates to the use of
tea juice as a raw material for obtaining valuable tea compounds such as
bioactive compounds and/or aroma compounds.

BACKGROUND TO THE INVENTION

[0002]Tea is a beverage traditionally made by infusing the dry leaves of
the plant Camellia sinensis in boiling water. Tea is (with the exception
of water) probably the world's most popular beverage and, in some parts
of the world, has traditionally been considered to have health-promoting
potential. Recently, extensive laboratory research and epidemiologic
studies have shown that many compounds present in tea show bioactivity
and may be useful, for example, in treating a variety of illnesses and/or
in producing enhanced physical or mental performance.

[0003]Polyphenolic compounds such as catechins and theaflavins have been
shown to be particularly valuable. Some of the benefits of tea
polyphenols may be directly linked to their antioxidant properties. The
purported benefits include lowering blood lipid levels (e.g.
cholesterol), anti-inflammation effects and anti-tumour effects.

[0004]Another tea compound which has been shown to have bioactivity is the
amino acid theanine. For example, it is reported that theanine stimulates
α-waves in the mammalian brain and bestows a relaxed but alert
mental state to the individual.

[0005]Besides bioactive compounds, tea also contains compounds which are
valued for their sensory qualities. In particular, tea has a unique aroma
and is rich in aroma compounds.

[0006]Although some of the benefits of tea compounds may be apparent at
consumption rates as low as a few cups per day, many individuals do not
even achieve this modest consumption rate on a long term basis.
Furthermore, tea beverages are less convenient to prepare than beverages
prepared from non-tea-based beverage precursors, such as instant coffee,
owing to the relatively slow rate of infusion of tea leaves and slow rate
of dissolution of tea powders. Also, there is an increasing desire
amongst consumers for products which deliver new sensory experiences but
which products are derived from natural sources.

[0007]There have therefore been many previous efforts to provide products
with enhanced levels of compounds derived from tea. In many cases the
previous efforts have employed a process wherein the tea compounds are
extracted from tea leaves using a solvent, such as water. For example, WO
2006/037511 (Unilever) discloses a process for preferentially extracting
theanine from tea plant material which involves a short cold water
extraction. One drawback with the known processes is that time and energy
are employed to remove the large amounts of solvent required for
exhaustive extraction.

[0008]Thus we have recognised that there is a need to provide a process
for obtaining tea compounds from tea leaf that does not require the use
of large amounts of a solvent. We have found that such a need can be met
by using the juice expressed from fresh tea leaves as a raw material for
the purification of tea compounds.

[0010]"Leaf tea" for the purposes of this invention means a tea product
that contains tea leaves and/or stem in an uninfused form, and that has
been dried to a moisture content of less than 30% by weight, and usually
has a water content in the range 1 to 10% by weight (i.e. "made tea").

[0012]"Fermentation" refers to the oxidative and hydrolytic process that
tea undergoes when certain endogenous enzymes and substrates are brought
together, e.g., by mechanical disruption of the cells by maceration of
the leaves. During this process colourless catechins in the leaves are
converted to a complex mixture of yellow and orange to dark-brown
polyphenolic substances.

[0013]"Fresh tea leaves" refers to tea leaves and/or stem that have never
been dried to a water content of less than 30% by weight, and usually
have a water content in the range 60 to 90%.

[0014]"Tea compound" refers to any compound derived from tea material
except for water. Thus tea compounds include all of the tea solids and
tea volatiles.

Expressing Juice

[0015]As used herein the term "expressing juice" refers to squeezing out
juice from fresh tea leaves using physical force, as opposed to
extraction of tea solids with the use of a solvent. Thus the term
"expressing" encompasses such means as squeezing, pressing, wringing,
spinning and extruding. It is possible that a small amount of solvent
(e.g. water) is added to the fresh leaves during the expression step.
However, in order to prevent significant extraction of tea solids by the
solvent, the moisture content of the leaves during expression is that of
fresh tea leaves as defined hereinabove. In other words, during the
expression step, the moisture content of the tea leaves is between 30 and
90% by weight, more preferably between 60 and 90%. It is also preferred
that the fresh leaves are not contacted with non-aqueous solvent (e.g.
alcohols) prior to or during expression, owing to the environmental &
economic problems associated with such solvents.

Polyphenol

[0016]As used herein, the term "polyphenol" refers to one or more of a
class compounds comprising a plurality of hydroxyl groups attached to one
or more aromatic groups. Typical tea polyphenols include catechin,
theaflavin and thearubigin.

[0017]As used herein the term "catechin" is used as a generic term for
catechin, gallocatechin, catechin gallate, gallocatechin gallate,
epicatechin, epigallocatechin, epicatechin gallate, epigallocatechin
gallate, and mixtures thereof.

[0018]As used herein the term "theaflavin" is used as a generic term for
theaflavin, isotheaflavin, neotheaflavin, theaflavin-3-gallate,
theaflavin-3'-gallate, theaflavin-3,3'-digallate, epitheaflavic acid,
epitheaflavic acid-3'-gallate, theaflavic acid, theaflavic
acid-3'-gallate and mixtures thereof. The structures of these compounds
are well-known (see, for example, structures xi-xx in Chapter 17 of
"Tea--Cultivation to consumption", K. C. Willson and M. N. Clifford
(Eds), 1992, Chapman & Hall, London, pp. 555-601). The term theaflavins
includes salt forms of these compounds. The preferred theaflavins are
theaflavin, theaflavin-3-gallate, theaflavin-3'-gallate,
theaflavin-3,3'-digallate and mixtures thereof, as these theaflavins are
most abundant in tea.

Beverage

[0019]As used herein the term "beverage" refers to a substantially aqueous
drinkable composition suitable for human consumption.

Leaf Size and Grade

[0020]For the purposes of the present invention, leaf particle size is
characterised by sieve mesh size using the following convention:
[0021]Tyler mesh sizes are used throughout. [0022]A "+" before the sieve
mesh indicates the particles are retained by the sieve. [0023]A "-"
before the sieve mesh indicates the particles pass through the sieve.

[0024]For example, if the particle size is described as -5 +20 mesh, then
the particles will pass through a 5 mesh sieve (particles smaller than
4.0 mm) and be retained by a 20 mesh sieve (particles larger than 841
μm).

[0025]Leaf particle size may additionally or alternatively be
characterized using the grades listed in the international standard ISO
6078-1982. These grades are discussed in detail in our European patent
specification EP 1 365 657 B1 (especially paragraph [0041] and Table 2)
which is hereby incorporated by reference.

Enrichment and Purification

[0026]Where a given composition is said to be "enriched" in a tea
compound, it is meant that the weight fraction of the tea compound in the
mixture of tea compounds in the composition is at least one and a half
times the weight fraction of the tea compound in the mixture of tea
compounds in the tea juice immediately following expression. This can be
expressed as shown in equation (1):

R=(cTC/cTOTAL)/(mTC/mTOTAL)≧1.5, (1)

wherein R is the enrichment factor of a particular tea compound in a given
composition, cTC is the mass of the particular tea compound in the
given composition, CTOTAL is the total mass of tea compounds in the
given composition, mTC is the mass of the particular tea compound in
the tea juice and mTOTAL is the total mass of tea compounds in the
tea juice.

[0027]Similarly "purification" refers to increasing the weight fraction of
a tea compound in a composition.

SUMMARY OF THE INVENTION

[0028]We have surprisingly found that juice expressed from tea leaves
contains a high content of valuable tea compounds whilst having a water
content lower than that typical of a conventional aqueous tea extracts.
Furthermore we have found that the juice can be readily fractionated to
yield compositions enriched in valuable tea compounds.

[0029]Thus the present invention provides a process comprising the steps
of: [0030]a) expressing juice from fresh tea leaves thereby to produce
leaf residue and juice comprising a mixture of tea compounds; [0031]b)
fractionating the mixture; and [0032]c) recovering at least one fraction
enriched in at least one tea compound.

[0033]In especially preferred embodiments, the at least one tea compound
is theanine and/or an aroma compound.

DETAILED DESCRIPTION

Expression of Juice

[0034]Step (a) of the process of the invention comprises expressing juice
from fresh tea leaves.

[0035]If the amount of juice expressed is too low then it becomes
difficult to separate the juice from the leaf residue and/or leads to an
inefficient process. Thus it is preferred that the amount of expressed
juice is at least 10 ml per kg of the fresh tea leaves, more preferably
at least 25 ml, more preferably still at least 50 ml and most preferably
from 75 to 600 ml. When referring to the volume of juice expressed per
unit mass of tea leaves it should be noted that the mass of the tea
leaves is expressed on an "as is" basis and not a dry weight basis. Thus
the mass includes any moisture in the leaves.

[0036]The expression step can be achieved in any convenient way so long as
it allows for separation of the tea juice from the leaf residue and
results in the required quantity of juice. The machinery used to express
the juice may, for example, include a hydraulic press, a pneumatic press,
a screw press, a belt press, an extruder or a combination thereof.

[0037]The juice may be obtained from the fresh leaves in a single pressing
or in multiple pressings of the fresh leaves. Preferably the juice is
obtained from a single pressing as this allows for a simple and rapid
process.

[0038]In order to minimise degradation of the valuable tea compounds, it
is preferred that the expression step is performed at ambient
temperature. For example, the leaf temperature may be from 5 to
40° C., more preferably 10 to 30° C.

[0039]The time and pressure used in the expression step can be varied to
yield the required amount of juice. Typically, however, the pressures
applied to express the juice will range from 0.5 MPa (73 psi) to 10 MPa
(1450 psi). The time over which the pressure is applied will typically
range from 1 s to 1 hour, more preferably from 10 s to 20 minutes and
most preferably from 30 s to 5 minutes.

[0040]Prior to expression, the fresh tea leaves may undergo a
pre-treatment including, for example, a unit process selected from heat
treatment to deactivate fermentation enzymes, maceration, withering,
fermentation or a combination thereof.

[0041]If the tea juice and/or leaf residue is to be used to obtain a green
tea compound (e.g. catechin) it is preferred that the fresh leaves are
heat treated to deactivate fermentation enzymes prior to expression.
Suitable heat treatments include steaming and/or pan-frying.

[0042]If the tea juice and/or leaf residue is to be used to obtain a black
or oolong tea compound (e.g. theaflavin and/or thearubigin) it is
preferred that the fresh leaves are not heat treated to deactivate
fermentation enzymes prior to expression. The fresh leaves may or may not
be fermented prior to expression. If the leaves are fermented prior to
expression then it is particularly preferred that they are macerated
prior to fermentation.

[0043]Whether or not the fresh leaves are fermented, maceration prior to
expression may help in decreasing the time and/or pressure required to
express the desired quantity of juice.

Fractionation

[0044]Step (b) of the process of the invention comprises fractionating the
mixture of tea compounds and step (c) comprises recovering at least one
fraction enriched in at least one tea compound.

[0045]The process may be used to purify any tea compound. However, the
preferred tea compounds are those which show bioactivity and/or
contribute to aroma. Thus it is preferred that the at least one tea
compound is polyphenol, amino acid or an aroma compound. Most preferably
the at least one tea compound is theanine and/or an aroma compound.

[0046]If the at least one tea compound is polyphenol, then it may be, for
example, catechin, theaflavin, thearubigin or a mixture thereof.

[0047]If the at least one tea compound is amino acid then it is preferably
theanine.

[0048]If the at least one tea compound is an aroma compound then it will
usually be volatile. By volatile is meant that it will have a vapour
pressure of at least 1 Pa at 25° C. Preferably the aroma compound
is methanol, acetaldehyde, dimethyl sulphide, 2-methyl-propanal, 2-methyl
butanal, 3-methyl butanal, 1-penten-3-one, hexanal, 1-penten-3-ol,
E-2-hexenal, Z-3-hexenyl acetate, Z-2-penten-1-ol, hexan-1-ol,
Z-3-hexenol, E-2-hexenol, cis-linalool oxide, 1-octen-3-ol,
trans-linalool oxide, linalool, α-terpinol, phenyl acetaldehyde,
methyl salicylate, geraniol, benzyl alcohol, 2-phenylethanol or a mixture
thereof.

[0049]Fractionation in step (b) can be achieved using any suitable process
capable of separating tea compounds. Examples of such processes include
the unit processes of membrane filtration, preparative chromatography,
solvent extraction, precipitation, distillation and combinations thereof.

[0050]Membrane filtration may include microfiltration, ultrafiltration,
nanofiltration, reverse osmosis or a combination thereof. The preferred
filtration operation comprises ultrafiltration, nanofiltration or a
combination thereof as these are especially effective at purifying
bioactive compounds such as polyphenol and/or amino acid. Typically
filtration will involve fractionating the mixture of tea compounds into
at least one permeate fraction and at least one retentate fraction.

[0051]As used herein, the term "preparative chromatography" refers to a
preparative process comprising the step of contacting the mixture of tea
compounds with a chromatographic medium. The chromatographic medium is a
substance which has a different affinity for at least 2 of the tea
compounds in the mixture, examples include adsorbant materials. Typically
the mixture will be fractionated by the preparative chromatography into
at least two fractions differing in the degree to which they interact
with the chromatographic medium. In a preferred embodiment, the
preparative chromatography is column chromatography. Where the
chromatography is column chromatography, the mixture will usually be
eluted from the column and fractions collected at varying elution times.

[0052]Solvent extraction preferably comprises contacting the mixture of
tea compounds with a solvent thereby to yield at least one soluble
fraction and at least one insoluble fraction.

[0053]Precipitation usually comprises subjecting the mixture to a physical
and/or chemical change such that soluble material precipitates out of
solution and/or suspended material sediments or creams. Examples of
chemical changes include changes in pH, solvent composition,
concentration or a combination thereof. Examples of physical changes
include heating or cooling, centrifugation or a combination thereof.

[0054]Distillation usually comprises heating the mixture to evaporate at
least some volatile tea compounds. It is especially preferred that step
(b) comprises distillation when the at least one tea compound is an aroma
compound.

[0055]The at least one fraction enriched in at least one tea compound
recovered on step (c) is preferably enriched in the tea compound such
that the enrichment factor R is at least 1.7, more preferably at least 2
and most preferably from 3 to 1000.

[0056]In a preferred embodiment the at least one fraction is concentrated
and/or dried. This allows for stable long-term storage of the fraction.
Typically the fraction will be dried to less than 20% moisture by weight,
more preferably less than 10% and optimally to 1 to 7% moisture.

[0057]It is preferred that the tea compounds from the tea juice are not
diluted with further tea compounds (e.g. from tea extract) prior to
fractionating the mixture as this will further complicate the
fractionation process.

Purification of Theanine

[0058]Tea juice is found to be especially rich in theanine compared with
conventional tea extracts. Thus in a preferred embodiment the process is
a process of purifying theanine from tea, the process comprising the
steps of: [0059]a) expressing juice from fresh tea leaves thereby to
produce leaf residue and juice comprising a mixture of tea compounds
comprising theanine; [0060]b) fractionating the mixture; and [0061]c)
recovering at least one fraction enriched in theanine.

[0062]In general, similar methods as those known for purifying theanine
from tea extracts can be employed in step (b). In particular, such
methods include nanofiltration (see, for example, WO 2006/037503) and/or
ion exclusion chromatography (see, for example, co-pending International
Patent Application No. PCT/EP2008/054817).

[0063]Preferably the at least one fraction enriched in theanine comprises
theanine in an amount of at least 5%, more preferably at least 8% and
most preferably from 10 to 100% by dry weight.

Purification of Aroma

[0064]Tea juice is a rich source of aroma compounds. Thus in a preferred
embodiment the process is a process of recovering aroma from tea, the
process comprising the steps of: [0065]a) expressing juice from fresh
tea leaves thereby to produce leaf residue and juice comprising a mixture
of tea compounds comprising aroma; [0066]b) fractionating the mixture;
and [0067]c) recovering at least one fraction enriched in aroma.

[0068]Preferably step (b) comprises a distillation step and step (c)
comprises recovering a distillate enriched in aroma. In general, similar
methods as those known for distilling aroma from tea extracts can be
employed in step (b). In particular, such methods include flash
evaporation (see, for example, WO 2003/101215) and/or carrier gas
distillation (see, for example, U.S. Pat. No. 4,880,656).

[0069]The at least one fraction enriched in aroma preferably has an aroma
content of at least 25 mg/l, more preferably at least 50 mg/l, more
preferably still at least 100 mg/l, and most preferably to an aroma
content in the range of 1000 mg/l to a concentrate that is purely aroma
oil (e.g. 900 g/l). The aroma content (or Total Organic Carbon--TOC--can
be determined by the method disclosed in WO 2007/079900).

Processing the Leaf Residue

[0070]In order to maximise the efficiency of the process it is preferred
that the leaf residue is not discarded but is further processed to
produce a commercially viable product. In a particularly preferred
embodiment, the process comprises an additional step (d) wherein the leaf
residue is processed to produce leaf tea.

[0071]We have surprisingly found that if the amount of juice expressed is
below 300 ml per kg of fresh leaves, the leaf residue can be processed to
make leaf tea of at least conventional quality despite the fact that the
leaf residue after expression has a lower overall level of tea compounds
such as polyphenols and amino acids. In general, the quality of the final
leaf tea (e.g. in terms of infusion performance) is better the less juice
expressed. Thus it is preferred that the amount of juice expressed in
step (a) is less than 300 ml per kg of tea leaves, more preferably less
than 275 ml, more preferably still less than 250 ml and most preferably
less than 225 ml.

[0074]A step common to manufacture of all leaf teas is a drying step. In
the case of oolong and black leaf tea, the drying step usually also
serves to deactivate the fermentation enzymes. Efficient drying requires
high temperatures and so it is preferred that step (d) of the process
comprises drying the leaf residue at a temperature of at least 75°
C., more preferably at least 90° C.

[0075]It is preferred that step (d) comprises sorting the leaf tea,
preferably after drying, to achieve a particle size of at least 35 mesh.
More preferably the leaf tea is sorted to achieve a particle size of from
30 mesh to 3 mesh. Alternatively or additionally, the leaf tea may be
sorted to achieve a leaf tea grade of Pekoe Fannings (PF) grade or
larger, more preferably Orange Fannings (OF) or larger and most
preferably Broken Orange Pekoe Fannings (BOPF) or larger.

EXAMPLES

[0076]The present invention will be further described with reference to
the following examples.

Example 1

[0077]This Example demonstrates the fractionation of tea juice using a
type of preparative chromatography known as solid phase extraction (SPE).

Collection of Juice

[0078]Fresh tea leaves (which had not been withered) were steamed for 60
seconds at ˜100° C. to inactivate endogenous enzymes and
thus prevent fermentation. Steamed leaves, cooled to room temperature,
were chopped using a vegetable cutter to yield chopped leaf of average
size of around 0.5 to 1 cm2. The dhool was then pressed using a
hydraulic press (5 Tonnes applied to a 500 g mass of leaf inside a
cylinder of diameter 160 mm, resulting in a downward pressure of 354 psi
(2.44 MPa)) to express green tea juice. The yield of green tea juice was
22 ml/100 g dhool, and had a total solids content of 8% by weight. The
tea juice was immediately centrifuged for 20 minutes (10000 g at
3° C.) and the supernatant was then filter-sterilised using a
Nalgene® filtration unit fitted with a 0.2 μm filter. The solids
content of the tea juice after centrifugation and filtration was 6% by
weight.

Fractionation of Juice

[0079]The green tea juice (1 ml) was applied to a C18 SPE cartridge
(SDB 200 mg/3 ml, purchased from J.T. Baker, Bakerbond SPE, Lot No.
0316910033). Following collection of the eluate, the cartridge was washed
with 1 ml water and then successive 1 ml volumes of methanol-water
mixtures (20%, 40%, 60%, 80% and 100% v/v methanol). An additional 4 ml
methanol wash was then applied to the cartridge column to ensure all of
the compounds adsorbed onto the column were completely washed off.
Catechins, caffeine and theanine were analysed from each fraction.

Results

[0080]The theanine, catechin and caffeine content of tea juice and various
methanol fractions is summarised in table 1. A small quantity of theanine
(yield of 7.7%) from the tea juice was found in the first eluent. All
caffeine and catechins were adsorbed by the SPE cartridge, as these were
not detected in the initial eluate. Water and 20% methanol fractions
recovered theanine in a yield of 60% and 27% respectively but neither
caffeine nor catechins were found in these two fractions. With the
increase of methanol in the mobile phase, catechins and caffeine started
to elute. Caffeine was mainly washed out when the percentage of methanol
increased up to 80%, while catechins mainly started to elute from 60% to
100% methanol washes. There are some interesting differences in elution
profiles between individual catechins. The 60% methanol wash eluted 73%
of EGC, while most of the ECG (61%) required 80% methanol to be eluted
from the column. EC and EGCG were mainly distributed in 60% and 80%
methanol fractions. Overall 1 ml of methanol was sufficient to completely
remove catechins from the column.

[0081]In a separate experiment, 1 ml tea juice was applied to each of six
SPE cartridges. The eluate from each cartridge was collected and bulked.
Each cartridge was then washed with 1 ml water and the resulting water
fractions bulked. Each cartridge was then washed with 1 ml 100% methanol
and the resulting methanol fractions bulked. The eluate, water and
methanol fractions were each freeze dried to a powder. In addition
unfractionated tea juice was directly freeze dried.

[0082]Theanine, caffeine and catechins were quantified in each powder by
ISO methods where applicable. Table 2 summarises the levels (mg/g dry
weight) of each component in each powder fraction.

[0086]The pressed residual dhool resulting from the juice production above
was broken up by hand and then dried using a fluidized bed drier (ten
minutes at 90° C., followed by ten minutes at 120° C.) to
obtain a leaf tea with moisture content of 3%.

Patent applications by David George Sharp, Sharnbrook GB

Patent applications by Steven Peter Colliver, Sharnbrook GB

Patent applications in class Of plant or plant derived material

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